Sialic acids (Sias) are a family of acidic sugars found on vertebrate glycoconjugates, being attached to underlying oligosaccharides (glycans) in different linkages, and modified in different ways. These linkages and modifications are molecule-specific, tissue-specific and developmentally regulated. The overall goal of this grant has been to unravel the mechanisms responsible for regulating some of these linkages and modifications, and to elucidate their biological and pathological roles. Thus, we have focused on understanding the biosynthetic mechanisms responsible for their expression, the specific intrinsic lectins that recognize them, the consequences for cellular signaling, differentiation, activation and survival, and for cellular interactions with the extracellular matrix. We are also interested in their roles in mediating cell-cell interactions and extracellular signaling during tissue organization and development, as well their recognition by specific lectins of extrinsic origin. In the past, this grant has supported studies of several such Sia modifications and linkages and the recognition of such modifications and linkages by various lectins. As our most recent interesting findings have been on the human-specific loss of the common mammalian Sia N-Glycolylneuraminic acid (Neu5Gc), we propose to focus exclusively on this issue during the next grant period. We hypothesize that this genetically determined loss has many implications, ranging from basic biochemical and cell biological effects involving uptake and incorporation into some cell types, to the presence and significance of anti-Neu5Gc antibodies, to an impact on the functions of both intrinsic and extrinsic receptors that recognize Sias. In addition to biochemical and cell biological studies, and studies of certain human samples, we will use mice with a human-like defect in Neu5Gc expression, and cell lines derived from such animals. This genetic approach increases the likelihood of definitive mechanistic conclusions to bolster the observational correlations in humans. Specific Aims are: #1, determining the fate of exogenously-introduced Neu5Gc in Neu5Gc-deficient Human cells and mice; #2, exploring the mechanism and consequences of incorporation of Neu5Gc from dietary sources, and the accompanying anti-Neu5Gc antibody response; #3, studying the consequences of human Neu5Gc loss on the evolution and function of intrinsic Sialic acid-recognizing receptors (particularly the CD33-related Siglecs); and, #4, exploring the consequences of human Neu5Gc loss on the evolution and function of extrinsic Sialic acid recognizing receptors. The extent to which some aspects of Aims #3 and #4 and their ramifications are pursued will depend on the results of #1 and #2, and on further preliminary studies. In the long run, these studies should help elucidate the molecular and cellular basis for certain diseases that appear to be uniquely or preferentially manifested in humans.